Abstract
Chitin is the second most abundant polysaccharide in nature after cellulose. The degradation products of chitin, amino-oligosaccharides, possess good biological activities and promising applications. Chitinase plays an important role in the degradation of chitin, and its highly efficient production has attracted wide attention. In this study, the Bacillus licheniformis-derived chitinase BlChiA was recombinantly expressed in Bacillus subtilis. The optimal promoter SpoVG and signal peptide YqxI were obtained by screening the endogenous promoters and signal peptides. The extracellular enzyme activity of the recombinant strain reached 1.96 U mL−1, 1.94-fold higher than that of the original, previously constructed strain. On this basis, mutant libraries were constructed using error-prone PCR, and mutants with enhanced activities were obtained using high-throughput activity assays. After combination, the mutant E336K/A472T exhibited 3.61-fold greater activity than the previous result, which represented the highest expression level of BlChiA in Bacillus subtilis. In addition, the mutant also exhibited greater acid adaptability. These results provided a foundation for further exploration of the function and industrial application of the enzyme.
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